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P3.052 Metallomics of neuromelanin in Parkinsonian syndromes
Poster presentations / Parkinsonism and Related Disorders 15S2 (2009) S29–S199
P3.049 b2 Adrenergic receptor activation induces microglial NADPH oxidase activation and dopaminergic neurotoxicity through an ERK-dependent/protein kinase A-independent pathway L. Qian1 , X. Hu2 , D. Zhang2 , B. Wilson2 , J.-S. Hong2 , P. Flood1 . 1 Comprehensive Center for Inflammatory Disorders, University of North Carolina at Chapel Hill, Chapel Hill, 2 Neuropharmocology, NIEHS, RTP, NC, USA Activation of the b2 adrenergic receptor (b2AR) on immune cells has been reported to possess anti-inflammatory properties, however, the pro-inflammatory properties of b2AR activation remain unclear. In this study, using rat primary mesencephalic neuron-glia cultures, we report that salmeterol, a long-acting b2AR agonist, selectively induces dopaminergic (DA) neurotoxicity through its ability to activate microglia. Salmeterol selectively increased the production of reactive oxygen species (ROS) by NADPH oxidase (PHOX), the superoxide-producing enzyme in microglia. A key role of PHOX in mediating salmeterol-induced neurotoxicity was demonstrated by the inhibition of DA neurotoxicity in cultures pretreated with diphenylene-iodonium (DPI), an inhibitor of PHOX activity. Mechanistic studies revealed the activation of microglia by salmeterol results in the selective phosphorylation of ERK, a signaling pathway required for the translocation of the PHOX cytosolic subunit p47phox to the cell membrane. Furthermore, we found ERK inhibition, but not protein kinase A (PKA) inhibition, significantly abolished salmeterol-induced superoxide production, p47phox translocation, and its ability to mediate neurotoxicity. Together, these findings indicate that b2AR activation induces microglial PHOX activation and DA neurotoxicity through an ERKdependent/PKA-independent pathway. P3.050 Role of tumor necrosis factor in the progressive phase of dopaminergic neuron death in a rat model of Parkinson’s disease A. Harms1 , S. Varghese1 , K. Ruhn1 , I. Trevino1 , A. Blesch2 , M. Tansey3 . 1 Physiology, UT Southwestern Medical Center, Dallas, TX, 2 Neuroscience, UCSD, San Diego, CA, 3 Physiology, Emory University School of Medicine, Atlanta, GA, USA TNF is involved in protection from bacterial infections, cell growth, viral replication, immune system regulation, and may have deleterious effects in autoimmune and neurodegenerative diseases. Chronic neuroinflammation is likely to be a key player in the progressive degeneration of the nigrostriatal system. Previous studies from our lab have shown that blocking soluble TNF signaling from the outset of a unilateral 6-hydroxydopamine (6-OHDA) striatal lesion attenuated behavioral deficits and dopaminergic neuron loss by 50% during the acute phase of cell death but did not address a possible role for TNF in the progressive loss of DA neurons, a critical question of clinical relevance to the human disease. Therefore, we determined the extent to which delayed inhibition of soluble TNF by a single intranigral injection of a lentivirus encoding a dominant negative TNF inhibitor afforded neuroprotection in the late and more progressive phases of 6-OHDA-induced cell death. Behavioral deficits were measured using sensitive sensorimotor tests and nigral DA neuron number was estimated by stereological analyses of NeuN/Tyrosine Hydroxylase-positive neurons in SNpc. Additional in vitro experiments are underway to elucidate the mechanism by which TNF contributes to the progressive loss of nigral DA neurons and the role of TNF signaling in regulating glial activation and survival responses of DA neurons. Funding provided by The Michael J. Fox Foundation for Parkinson’s Research and NINDS 5R01NS049433-03.
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P3.051 Calcium channel blockers can minimize iron-induced apoptosis in neural stem cells W.-X. Chen, V. Wong, M. Yang, G.C.-F. Chan. Pediatrics and Adolescent Medicine, The University of Hong Kong, Hong Kong, Hong Kong S.A.R. Iron accumulation in the brain with oxidative damage has commonly been observed in hemorrhagic stroke and neurodegenerative disorders. L-type calcium channels are key passages for iron entry into neuronal cells. We postulated calcium channel blockers (CCBs) could protect against iron-induced neurotoxicity in neural stem cells (NSCs). In vitro studies of iron overload on murine-derived neural stem cell line C17.2 and rat embryonic (E13.5) hippocampal neural stem cells were performed. The cytotoxic effect of iron (ferrous ammonium sulfate, ferric chloride and ferric ammonium citrate) on NSCs and the salvaging potential of CCBs (nimodipine and flunarizine) were studied. Cell viability was measured by XTT assay. Apoptotic cell death was assessed by annexin V/PI staining, activated caspase-3 and mitochondrial membrane potential (JC-1) using flow cytometry. The effect of iron overload on ERK activation as well as the rescuing effect of CCBs and MEK inhibitor (U0126) on ERK phosphorylation of iron-injured NSCs was explored by flow cytometry. Results: Iron overload significantly decreased cell viability via inducing mitochondrial damage and caspase-3 activity in a doseand time-dependent manner (0.15–1.8 mM, 24 & 48 hrs). Clinical relevant doses of iron (0.6–0.9 mM) induced ERK activation of NSCs. Under such condition, CCBs could alleviate ERK activation and significantly improve cell viability by partially ameliorated apoptosis via preventing mitochondrial damage and caspase-3 activity. U0126 prevented ERK activation and abrogated apoptosis in iron-overloaded NSCs. Conclusions: Iron-induced apoptosis of NSCs underwent mitochondria-mediated apoptotic pathway and it involved ERK activation. CCBs could potentially protect iron-induced neurotoxicity in NSCs by inhibiting these processes. P3.052 Metallomics of neuromelanin in Parkinsonian syndromes S. Bohic1 , A. Carmona2 , R. Ortega2 , S. Reyes3 , F. Carew-Jones3 , K. Double3 . 1 INSERM U-836 (Equipe 6) Rayonnement Synchrotron et Recherche M´edicale, Grenoble Institut des Neurosciences, Grenoble, 2 CNAB UMR5084, CENBG, Gradignan, France; 3 Prince of Wales Medical Research Institute and the University of New South Wales, Sydney, NSW, Australia We investigated changes in levels of transition metals within brain tissue, suggested to contribute to neurodegenerative cascades in Parkinson’s disease (PD), using highly sensitive, high resolution methods. Twenty micron formalin-fixed sections of the substantia nigra (SN) were prepared from five controls, five idiopathic PD, three Incidental Lewy body disease (ILBD) and three Alzheimer’s disease (AD) brains. X-ray absorption microspectroscopy (microXANES) determination of sulphur oxidation state within NM was performed and a semi-quantitative description of the spectra was estimated by optimizing a linear combination of the set of known reference compounds from different sulphur species. NM-metal content was measured in 6–7 individual pigmented cells per case using synchrotron X-ray fluorescence and in a 600×600 micron area of SN tissues using particle-induced X-ray emission (PIXE). The sulphur environment of the pigment polymer and concentrations of NM-associated calcium, manganese, iron, zinc and selenium did not differ between diagnostic groups. In contrast, using both X-ray fluorescence and PIXE on independent tissue sections, NMassociated copper (Cu) was found to be significantly decreased in PD and ILBD, but not in AD. PIXE also demonstrated a significant decrease in non-melanised tissue Cu in PD and ILBD. This work reveals a novel decrease in NM- and tissue-associated Cu content
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Poster presentations / Parkinsonism and Related Disorders 15S2 (2009) S29–S199
in PD. Decreased Cu was also seen in ILBD, a disorder suggested to represent preclinical PD, suggesting this change occurs early in these synucleinopathies. Our findings may be relevant for Cucontaining proteins, such as superoxide dismutase and alphasynuclein, thought to be involved in degenerative mechanisms in parkinsonian syndromes. P3.053 Up-regulation of divalent metal transporter 1 in 6-hydroxydopamine intoxication is IRE/IRP dependent H. Jiang, N. Song, L. Hou, H. Xu, S. Zhang, J. Xie. State Key Disciplines: Physiology (in incubation); Department of Physiology, Qingdao University, Qingdao, China Iron plays a key role in Parkinson’s disease (PD) and increased iron content was found in the substantia nigra (SN) in PD patients. Divalent metal transporter 1 (DMT1) was up-regulated in the SN of both MPTP-induced PD models and PD patients. However, what are the mechanisms underlying DMT1 up-regulation is largely unknown. In the present study, we first observed DMT1 with the iron responsive element (IRE, DMT1+IRE) but not DMT1 without IRE (DMT1−IRE) was up-regulated in vivo in the SN of 6-hydroxydopamine (6-OHDA)-induced PD rats, suggesting increased DMT1+IRE expression might account for the nigral iron accumulation in PD rats. This was futher confirmed by our in vitro study in 6-OHDA-treated and DMT1+IRE overexpressed MES23.5 cells, respectively. Increased iron regulatory protein (IRP) 1 and IRP2 expression were observed, while silencing of IRPs dramatically diminished 6-OHDA-induced DMT1+IRE upregulation. Pretreatment with N-acetyl-L-cysteine could fully suppress IRPs up-regulations by inhibiting 6-OHDA-induced oxidative stress. Our data gave the direct evidence that DMT1+IRE up-regulation accounted for 6-OHDA-induced iron accumulation in an IRE/IRP-dependent manner, which was initiated by 6-OHDAinduced intracellular oxidative stress. Increased intracellular iron thus resulted in aggravated oxidative stress. This study might support the use of anti-oxidants in the treatment of PD from a quite new angel, which could inhibit iron accumulation by regulating DMT1 expression. P3.054 Effect of prenatal manganese intoxication on dopaminergic neurotranssmision in the brain of rats lesioned as neonates with 6-hydroxydopamine 1
1
2
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P. Nowak , A. Kwiecinski , R.M. Kostrzewa . Pharmacology, University, Zabrze, Poland; 2 Pharmacology, University, Johnson City, TN, USA The effects of prenatal manganese (Mn) intoxication on reactivity of central dopamine receptors and dopamine (DA) metabolism were studied in rats lesioned as neonates with 6-hydroxydopamine (6-OHDA). MnCl2 × 4H2 O (10,000 ppm) was added to the drinking water of Wistar pregnant rats for the duration of pregnancy. On the day of parturition Mn was discontinued as an additive in the drinking water. The control group consisted of rats that consumed water without Mn. At 3 days after birth rats of both examined groups (control and Mn) were pretreated with desipramine HCl (20 mg/kg) and pargyline HCl (50 mg/kg) and injected bilaterally icv with 6-OHDA HBr in one of three doses (15 mg, 30 mg or 67 mg base form on each side, or the vehicle saline) (control). At 8 weeks after birth rats were injected with either SKF 38393 (DA D1 agonist), apomorphine (DA D2 agonist), quinpirole (DA D2/D3 agonist), SCH 23390 (DA D1 antagonist) or haloperidol (DA D2 antagonist), and were tested for characteristic behavioral effects: oral movements, yawning, stereotypy and catalepsy. Furthermore DA and its metabolites as well as DA-synthesis rate in the frontal cortex, hippocampus, striatum, hypothalamus, cerebellum and brain stem were evaluated. It was shown that Mn intoxication produced disturbances mainly in
corticolimbic DA neurotransmission. Moreover based on behavioral data we demonstrated that Mn increased the vulnerability to neurotoxic effects of the 15 mg dose of 6-OHDA. P3.055 Welding fume-related dopaminergic neurotoxicity K. Sriram, G.X. Lin, A.M. Jefferson, O. Wirth, Y. Hayashi, J.R. Roberts, R.S. Chapman, K.M. Krajnak, J.M. Antonini. CDC-NIOSH, Morgantown, WV, USA Manganese (Mn) intoxication is associated with increased prevalence of neurological conditions resembling Parkinson’s disease (PD). The presence of Mn in welding consumables therefore raises concerns about potential neurological risks following occupational exposure to welding fume (WF). To investigate if WF exposure induces neurotoxicity, we treated rats with low- and high-Mn containing WF. Repeated intratracheal instillations (0.5 mg/rat, 1/wk × 7 wks) of gas metal arc-mild steel (GMA-MS; low Mn, insoluble) or manual metal arc-hard surfacing (MMA-HS; high Mn, soluble) fumes caused loss of tyrosine hydroxylase (TH) protein in the striatum (~20%) and midbrain (~30%) by 1 d post-exposure. While loss of TH following GMA-MS was transient, a sustained loss was observed in the midbrain (34%) even after cessation of MMA-HS exposure. Both fumes caused down-regulation of D2 receptor (30–40%) and Vmat2 (30–55%) mRNAs in the midbrain. A different exposure paradigm (2 mg/rat, 1/wk × 28 wks) resulted in loss of Park 5 and Park 7 proteins in striatum (40–50%), but not midbrain. A similar regimen of MnCl2 caused extensive loss of these proteins in both striatum (70–80%) and midbrain (60%). All treated animals exhibited hyperactivity but not deficits in gross-motor behavior. As mutations in PARK genes have been linked to onset of PD and because welding is implicated as a risk factor for idiopathic PD, it remains to be elucidated if PARK genes have an underlying role in the pathogenesis of WF-induced neurodegeneration. Our findings suggest that Mn-containing WF can cause persistent alteration of several PD-related genes that could lead to dopaminergic neurodegeneration. P3.056 The relationship between blood levels of heavy metals and Parkinson’s disease in China X. Tan. School of Public Health, Wuhan University, Wuhan, China Background: It is well known that idiopathic Parkinson’s disease (PD) and the Parkinsonian syndrome caused by manganese (Mn) poisoning are undistinguishable in the symptoms. So we doubt there are many patients who have accumulated Mn in the body in daily life and are diagnosed as idiopathic PD. We examined the relationship between PD and blood levels of heavy metals, and the influence of their intake from food in a population in general environment. Methods: The subjects comprising PD patients and sex- and agematched controls were recruited from the outpatient clinic of Xiangfan No. 1 People’s Hospital in Hubei, China between 2006 and 2008. Previous eating habits etc. before they were diagnosed with PD were asked. The morning blood samples were collected before breakfast and were used to measure concentrations of metals. Results: Whole blood Mn and serum iron (Fe) levels were significantly higher in PD patients than in controls. But no differences were seen in intake of each metal from food, experience of Mn poisoning between groups. Conclusions: In China, accumulation of Mn and Fe from unknown route might be involved in the etiology of PD in a general population.
P3.049 b2 Adrenergic receptor activation induces microglial NADPH oxidase activation and dopaminergic neurotoxicity through an ERK-dependent/protein kinase A-independent pathway L. Qian1 , X. Hu2 , D. Zhang2 , B. Wilson2 , J.-S. Hong2 , P. Flood1 . 1 Comprehensive Center for Inflammatory Disorders, University of North Carolina at Chapel Hill, Chapel Hill, 2 Neuropharmocology, NIEHS, RTP, NC, USA Activation of the b2 adrenergic receptor (b2AR) on immune cells has been reported to possess anti-inflammatory properties, however, the pro-inflammatory properties of b2AR activation remain unclear. In this study, using rat primary mesencephalic neuron-glia cultures, we report that salmeterol, a long-acting b2AR agonist, selectively induces dopaminergic (DA) neurotoxicity through its ability to activate microglia. Salmeterol selectively increased the production of reactive oxygen species (ROS) by NADPH oxidase (PHOX), the superoxide-producing enzyme in microglia. A key role of PHOX in mediating salmeterol-induced neurotoxicity was demonstrated by the inhibition of DA neurotoxicity in cultures pretreated with diphenylene-iodonium (DPI), an inhibitor of PHOX activity. Mechanistic studies revealed the activation of microglia by salmeterol results in the selective phosphorylation of ERK, a signaling pathway required for the translocation of the PHOX cytosolic subunit p47phox to the cell membrane. Furthermore, we found ERK inhibition, but not protein kinase A (PKA) inhibition, significantly abolished salmeterol-induced superoxide production, p47phox translocation, and its ability to mediate neurotoxicity. Together, these findings indicate that b2AR activation induces microglial PHOX activation and DA neurotoxicity through an ERKdependent/PKA-independent pathway. P3.050 Role of tumor necrosis factor in the progressive phase of dopaminergic neuron death in a rat model of Parkinson’s disease A. Harms1 , S. Varghese1 , K. Ruhn1 , I. Trevino1 , A. Blesch2 , M. Tansey3 . 1 Physiology, UT Southwestern Medical Center, Dallas, TX, 2 Neuroscience, UCSD, San Diego, CA, 3 Physiology, Emory University School of Medicine, Atlanta, GA, USA TNF is involved in protection from bacterial infections, cell growth, viral replication, immune system regulation, and may have deleterious effects in autoimmune and neurodegenerative diseases. Chronic neuroinflammation is likely to be a key player in the progressive degeneration of the nigrostriatal system. Previous studies from our lab have shown that blocking soluble TNF signaling from the outset of a unilateral 6-hydroxydopamine (6-OHDA) striatal lesion attenuated behavioral deficits and dopaminergic neuron loss by 50% during the acute phase of cell death but did not address a possible role for TNF in the progressive loss of DA neurons, a critical question of clinical relevance to the human disease. Therefore, we determined the extent to which delayed inhibition of soluble TNF by a single intranigral injection of a lentivirus encoding a dominant negative TNF inhibitor afforded neuroprotection in the late and more progressive phases of 6-OHDA-induced cell death. Behavioral deficits were measured using sensitive sensorimotor tests and nigral DA neuron number was estimated by stereological analyses of NeuN/Tyrosine Hydroxylase-positive neurons in SNpc. Additional in vitro experiments are underway to elucidate the mechanism by which TNF contributes to the progressive loss of nigral DA neurons and the role of TNF signaling in regulating glial activation and survival responses of DA neurons. Funding provided by The Michael J. Fox Foundation for Parkinson’s Research and NINDS 5R01NS049433-03.
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P3.051 Calcium channel blockers can minimize iron-induced apoptosis in neural stem cells W.-X. Chen, V. Wong, M. Yang, G.C.-F. Chan. Pediatrics and Adolescent Medicine, The University of Hong Kong, Hong Kong, Hong Kong S.A.R. Iron accumulation in the brain with oxidative damage has commonly been observed in hemorrhagic stroke and neurodegenerative disorders. L-type calcium channels are key passages for iron entry into neuronal cells. We postulated calcium channel blockers (CCBs) could protect against iron-induced neurotoxicity in neural stem cells (NSCs). In vitro studies of iron overload on murine-derived neural stem cell line C17.2 and rat embryonic (E13.5) hippocampal neural stem cells were performed. The cytotoxic effect of iron (ferrous ammonium sulfate, ferric chloride and ferric ammonium citrate) on NSCs and the salvaging potential of CCBs (nimodipine and flunarizine) were studied. Cell viability was measured by XTT assay. Apoptotic cell death was assessed by annexin V/PI staining, activated caspase-3 and mitochondrial membrane potential (JC-1) using flow cytometry. The effect of iron overload on ERK activation as well as the rescuing effect of CCBs and MEK inhibitor (U0126) on ERK phosphorylation of iron-injured NSCs was explored by flow cytometry. Results: Iron overload significantly decreased cell viability via inducing mitochondrial damage and caspase-3 activity in a doseand time-dependent manner (0.15–1.8 mM, 24 & 48 hrs). Clinical relevant doses of iron (0.6–0.9 mM) induced ERK activation of NSCs. Under such condition, CCBs could alleviate ERK activation and significantly improve cell viability by partially ameliorated apoptosis via preventing mitochondrial damage and caspase-3 activity. U0126 prevented ERK activation and abrogated apoptosis in iron-overloaded NSCs. Conclusions: Iron-induced apoptosis of NSCs underwent mitochondria-mediated apoptotic pathway and it involved ERK activation. CCBs could potentially protect iron-induced neurotoxicity in NSCs by inhibiting these processes. P3.052 Metallomics of neuromelanin in Parkinsonian syndromes S. Bohic1 , A. Carmona2 , R. Ortega2 , S. Reyes3 , F. Carew-Jones3 , K. Double3 . 1 INSERM U-836 (Equipe 6) Rayonnement Synchrotron et Recherche M´edicale, Grenoble Institut des Neurosciences, Grenoble, 2 CNAB UMR5084, CENBG, Gradignan, France; 3 Prince of Wales Medical Research Institute and the University of New South Wales, Sydney, NSW, Australia We investigated changes in levels of transition metals within brain tissue, suggested to contribute to neurodegenerative cascades in Parkinson’s disease (PD), using highly sensitive, high resolution methods. Twenty micron formalin-fixed sections of the substantia nigra (SN) were prepared from five controls, five idiopathic PD, three Incidental Lewy body disease (ILBD) and three Alzheimer’s disease (AD) brains. X-ray absorption microspectroscopy (microXANES) determination of sulphur oxidation state within NM was performed and a semi-quantitative description of the spectra was estimated by optimizing a linear combination of the set of known reference compounds from different sulphur species. NM-metal content was measured in 6–7 individual pigmented cells per case using synchrotron X-ray fluorescence and in a 600×600 micron area of SN tissues using particle-induced X-ray emission (PIXE). The sulphur environment of the pigment polymer and concentrations of NM-associated calcium, manganese, iron, zinc and selenium did not differ between diagnostic groups. In contrast, using both X-ray fluorescence and PIXE on independent tissue sections, NMassociated copper (Cu) was found to be significantly decreased in PD and ILBD, but not in AD. PIXE also demonstrated a significant decrease in non-melanised tissue Cu in PD and ILBD. This work reveals a novel decrease in NM- and tissue-associated Cu content
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Poster presentations / Parkinsonism and Related Disorders 15S2 (2009) S29–S199
in PD. Decreased Cu was also seen in ILBD, a disorder suggested to represent preclinical PD, suggesting this change occurs early in these synucleinopathies. Our findings may be relevant for Cucontaining proteins, such as superoxide dismutase and alphasynuclein, thought to be involved in degenerative mechanisms in parkinsonian syndromes. P3.053 Up-regulation of divalent metal transporter 1 in 6-hydroxydopamine intoxication is IRE/IRP dependent H. Jiang, N. Song, L. Hou, H. Xu, S. Zhang, J. Xie. State Key Disciplines: Physiology (in incubation); Department of Physiology, Qingdao University, Qingdao, China Iron plays a key role in Parkinson’s disease (PD) and increased iron content was found in the substantia nigra (SN) in PD patients. Divalent metal transporter 1 (DMT1) was up-regulated in the SN of both MPTP-induced PD models and PD patients. However, what are the mechanisms underlying DMT1 up-regulation is largely unknown. In the present study, we first observed DMT1 with the iron responsive element (IRE, DMT1+IRE) but not DMT1 without IRE (DMT1−IRE) was up-regulated in vivo in the SN of 6-hydroxydopamine (6-OHDA)-induced PD rats, suggesting increased DMT1+IRE expression might account for the nigral iron accumulation in PD rats. This was futher confirmed by our in vitro study in 6-OHDA-treated and DMT1+IRE overexpressed MES23.5 cells, respectively. Increased iron regulatory protein (IRP) 1 and IRP2 expression were observed, while silencing of IRPs dramatically diminished 6-OHDA-induced DMT1+IRE upregulation. Pretreatment with N-acetyl-L-cysteine could fully suppress IRPs up-regulations by inhibiting 6-OHDA-induced oxidative stress. Our data gave the direct evidence that DMT1+IRE up-regulation accounted for 6-OHDA-induced iron accumulation in an IRE/IRP-dependent manner, which was initiated by 6-OHDAinduced intracellular oxidative stress. Increased intracellular iron thus resulted in aggravated oxidative stress. This study might support the use of anti-oxidants in the treatment of PD from a quite new angel, which could inhibit iron accumulation by regulating DMT1 expression. P3.054 Effect of prenatal manganese intoxication on dopaminergic neurotranssmision in the brain of rats lesioned as neonates with 6-hydroxydopamine 1
1
2
1
P. Nowak , A. Kwiecinski , R.M. Kostrzewa . Pharmacology, University, Zabrze, Poland; 2 Pharmacology, University, Johnson City, TN, USA The effects of prenatal manganese (Mn) intoxication on reactivity of central dopamine receptors and dopamine (DA) metabolism were studied in rats lesioned as neonates with 6-hydroxydopamine (6-OHDA). MnCl2 × 4H2 O (10,000 ppm) was added to the drinking water of Wistar pregnant rats for the duration of pregnancy. On the day of parturition Mn was discontinued as an additive in the drinking water. The control group consisted of rats that consumed water without Mn. At 3 days after birth rats of both examined groups (control and Mn) were pretreated with desipramine HCl (20 mg/kg) and pargyline HCl (50 mg/kg) and injected bilaterally icv with 6-OHDA HBr in one of three doses (15 mg, 30 mg or 67 mg base form on each side, or the vehicle saline) (control). At 8 weeks after birth rats were injected with either SKF 38393 (DA D1 agonist), apomorphine (DA D2 agonist), quinpirole (DA D2/D3 agonist), SCH 23390 (DA D1 antagonist) or haloperidol (DA D2 antagonist), and were tested for characteristic behavioral effects: oral movements, yawning, stereotypy and catalepsy. Furthermore DA and its metabolites as well as DA-synthesis rate in the frontal cortex, hippocampus, striatum, hypothalamus, cerebellum and brain stem were evaluated. It was shown that Mn intoxication produced disturbances mainly in
corticolimbic DA neurotransmission. Moreover based on behavioral data we demonstrated that Mn increased the vulnerability to neurotoxic effects of the 15 mg dose of 6-OHDA. P3.055 Welding fume-related dopaminergic neurotoxicity K. Sriram, G.X. Lin, A.M. Jefferson, O. Wirth, Y. Hayashi, J.R. Roberts, R.S. Chapman, K.M. Krajnak, J.M. Antonini. CDC-NIOSH, Morgantown, WV, USA Manganese (Mn) intoxication is associated with increased prevalence of neurological conditions resembling Parkinson’s disease (PD). The presence of Mn in welding consumables therefore raises concerns about potential neurological risks following occupational exposure to welding fume (WF). To investigate if WF exposure induces neurotoxicity, we treated rats with low- and high-Mn containing WF. Repeated intratracheal instillations (0.5 mg/rat, 1/wk × 7 wks) of gas metal arc-mild steel (GMA-MS; low Mn, insoluble) or manual metal arc-hard surfacing (MMA-HS; high Mn, soluble) fumes caused loss of tyrosine hydroxylase (TH) protein in the striatum (~20%) and midbrain (~30%) by 1 d post-exposure. While loss of TH following GMA-MS was transient, a sustained loss was observed in the midbrain (34%) even after cessation of MMA-HS exposure. Both fumes caused down-regulation of D2 receptor (30–40%) and Vmat2 (30–55%) mRNAs in the midbrain. A different exposure paradigm (2 mg/rat, 1/wk × 28 wks) resulted in loss of Park 5 and Park 7 proteins in striatum (40–50%), but not midbrain. A similar regimen of MnCl2 caused extensive loss of these proteins in both striatum (70–80%) and midbrain (60%). All treated animals exhibited hyperactivity but not deficits in gross-motor behavior. As mutations in PARK genes have been linked to onset of PD and because welding is implicated as a risk factor for idiopathic PD, it remains to be elucidated if PARK genes have an underlying role in the pathogenesis of WF-induced neurodegeneration. Our findings suggest that Mn-containing WF can cause persistent alteration of several PD-related genes that could lead to dopaminergic neurodegeneration. P3.056 The relationship between blood levels of heavy metals and Parkinson’s disease in China X. Tan. School of Public Health, Wuhan University, Wuhan, China Background: It is well known that idiopathic Parkinson’s disease (PD) and the Parkinsonian syndrome caused by manganese (Mn) poisoning are undistinguishable in the symptoms. So we doubt there are many patients who have accumulated Mn in the body in daily life and are diagnosed as idiopathic PD. We examined the relationship between PD and blood levels of heavy metals, and the influence of their intake from food in a population in general environment. Methods: The subjects comprising PD patients and sex- and agematched controls were recruited from the outpatient clinic of Xiangfan No. 1 People’s Hospital in Hubei, China between 2006 and 2008. Previous eating habits etc. before they were diagnosed with PD were asked. The morning blood samples were collected before breakfast and were used to measure concentrations of metals. Results: Whole blood Mn and serum iron (Fe) levels were significantly higher in PD patients than in controls. But no differences were seen in intake of each metal from food, experience of Mn poisoning between groups. Conclusions: In China, accumulation of Mn and Fe from unknown route might be involved in the etiology of PD in a general population.